SummaryThe Aryl hydrocarbon receptor is an evolutionary conserved widely expressed transcription factor that mediates the toxicity of a substantial variety of exogenous toxins, but is also stimulated by endogenous physiological ligands. While it is known that this receptor mediates the toxicity of dioxin, this is unlikely to be its physiological function. We have recently identified selective expression of AhR in the Th17 subset of effector CD4 T cells. Ligation of AhR by a candidate endogenous ligand (FICZ) which is a UV metabolite of tryptophan causes expansion of Th17 cells and the induction of IL-22 production. As a consequence, AhR ligation will exacerbate autoimmune diseases such as experimental autoimmune encephalomyelitis. Little is known so far about the impact of AhR ligands on IL-17/IL-22 mediated immune defense functions. IL-22 is considered a pro-inflammatory Th17 cytokine, which is involved in the etiology of psoriasis, but it has also been shown to be a survival factor for epithelial cells. AhR is polymorphic and defined as high or low affinity receptor for dioxin leading to the classification of high and low responder mouse strains based on defined mutations. In humans similar polymorphisms exist and although on the whole human AhR is thought to be of low affinity in humans, there are identified mutations that confer high responder status. No correlations have been made with Th17 mediated immune responses in mice and humans. This study aims to investigate the role of AhR ligands and polymorphisms in autoimmunity as well as protective immune responses using both mouse models and human samples from normal controls as well as psoriasis patients.

The Aryl hydrocarbon receptor is an evolutionary conserved widely expressed transcription factor that mediates the toxicity of a substantial variety of exogenous toxins, but is also stimulated by endogenous physiological ligands. While it is known that this receptor mediates the toxicity of dioxin, this is unlikely to be its physiological function. We have recently identified selective expression of AhR in the Th17 subset of effector CD4 T cells. Ligation of AhR by a candidate endogenous ligand (FICZ) which is a UV metabolite of tryptophan causes expansion of Th17 cells and the induction of IL-22 production. As a consequence, AhR ligation will exacerbate autoimmune diseases such as experimental autoimmune encephalomyelitis. Little is known so far about the impact of AhR ligands on IL-17/IL-22 mediated immune defense functions. IL-22 is considered a pro-inflammatory Th17 cytokine, which is involved in the etiology of psoriasis, but it has also been shown to be a survival factor for epithelial cells. AhR is polymorphic and defined as high or low affinity receptor for dioxin leading to the classification of high and low responder mouse strains based on defined mutations. In humans similar polymorphisms exist and although on the whole human AhR is thought to be of low affinity in humans, there are identified mutations that confer high responder status. No correlations have been made with Th17 mediated immune responses in mice and humans. This study aims to investigate the role of AhR ligands and polymorphisms in autoimmunity as well as protective immune responses using both mouse models and human samples from normal controls as well as psoriasis patients.

SummaryThis project focuses on two questions about host/parasite interactions: how do biotrophic plant pathogens suppress host defence? and, what is the basis for pathogen specialization on specific host species? A broadly accepted model explains resistance and susceptibility to plant pathogens. First, pathogens make conserved molecules ( PAMPS ) such as flagellin, that plants detect via cell surface receptors, leading to PAMP-Triggered Immunity (PTI). Second, pathogens make effectors that suppress PTI. Third, plants carry 100s of Resistance (R) genes that detect an effector, and activate Effector-Triggered Immunity (ETI). One effector is sufficient to trigger resistance. Albugo candida (Ac) (white rust) strongly suppresses host defence; Ac-infected Arabidopsis are susceptible to pathogen races to which they are otherwise resistant. Ac is an oomycete, not a fungus. Arabidopsis is resistant to races of Ac that infect brassicas. The proposed project involves three programs. First ( genomics, transcriptomics and bioinformatics ), we will use next-generation sequencing (NGS) methods (Solexa and GS-Flex), and novel transcriptomics methods to define the genome sequence and effector set of three Ac strains, as well as carrying out >40- deep resequencing of 7 additional Ac strains. Second, ( effectoromics ), we will carry out functional assays using Effector Detector Vectors (Sohn Plant Cell 19:4077 [2007]), with the set of Ac effectors, screening for enhanced virulence, for suppression of defence, for effectors that are recognized by R genes in disease resistant Arabidopsis and for host effector targets. Third, ( resistance diversity ), we will characterize Arabidopsis germplasm for R genes to Ac, both for recognition of Arabidopsis strains of Ac, and for recognition in Arabidopsis of effectors from Ac strains that infect brassica. This proposal focuses on Ac, but will establish methods that could discover new R genes in non-hosts against many plant diseases.

This project focuses on two questions about host/parasite interactions: how do biotrophic plant pathogens suppress host defence? and, what is the basis for pathogen specialization on specific host species? A broadly accepted model explains resistance and susceptibility to plant pathogens. First, pathogens make conserved molecules ( PAMPS ) such as flagellin, that plants detect via cell surface receptors, leading to PAMP-Triggered Immunity (PTI). Second, pathogens make effectors that suppress PTI. Third, plants carry 100s of Resistance (R) genes that detect an effector, and activate Effector-Triggered Immunity (ETI). One effector is sufficient to trigger resistance. Albugo candida (Ac) (white rust) strongly suppresses host defence; Ac-infected Arabidopsis are susceptible to pathogen races to which they are otherwise resistant. Ac is an oomycete, not a fungus. Arabidopsis is resistant to races of Ac that infect brassicas. The proposed project involves three programs. First ( genomics, transcriptomics and bioinformatics ), we will use next-generation sequencing (NGS) methods (Solexa and GS-Flex), and novel transcriptomics methods to define the genome sequence and effector set of three Ac strains, as well as carrying out >40- deep resequencing of 7 additional Ac strains. Second, ( effectoromics ), we will carry out functional assays using Effector Detector Vectors (Sohn Plant Cell 19:4077 [2007]), with the set of Ac effectors, screening for enhanced virulence, for suppression of defence, for effectors that are recognized by R genes in disease resistant Arabidopsis and for host effector targets. Third, ( resistance diversity ), we will characterize Arabidopsis germplasm for R genes to Ac, both for recognition of Arabidopsis strains of Ac, and for recognition in Arabidopsis of effectors from Ac strains that infect brassica. This proposal focuses on Ac, but will establish methods that could discover new R genes in non-hosts against many plant diseases.

Max ERC Funding

2 498 923 €

Duration

Start date: 2009-01-01, End date: 2014-06-30

Project acronymDCPOIESIS

ProjectSteady-state and demand-driven dendritic cell generation

Researcher (PI)Caetano Maria Pacheco Pais Dos Reis e Sousa

Host Institution (HI)THE FRANCIS CRICK INSTITUTE LIMITED

Call DetailsAdvanced Grant (AdG), LS6, ERC-2017-ADG

SummaryClassical dendritic cells (cDCs) are leucocytes that play a key role in innate immunity as well as the initiation and regulation of T cell responses. cDCpoiesis starts with commitment of a bone marrow (BM) haematopoietic progenitor, known as the classical DC precursor (CDP), to the cDC lineage. CDPs then give rise to pre-cDCs that exit the BM via the blood and seed tissues to give rise to the two major types of fully-differentiated cDCs, the cDC1 and cDC2 subsets. The key parameters of cDCpoiesis are poorly understood. We propose to characterise the niche in which cDCs develop within the BM and to study how pre-cDCs seed tissues and establish local clones of differentiated cDC1 and cDC2. We further wish to ask how the activity of CDPs and pre-cDCs is altered following infection, inflammation or tissue damage. Finally, we want to know to what extent cDCpoiesis is affected by direct sensing of infection or cell damage by cDC precursors. All these objectives will be addressed in a mouse lineage tracing model in which cDC precursors are genetically labelled through the activity of a Cre recombinase driven by the Clec9a locus. These mice will be crossed to fluorescent protein reporter mice, including Confetti mice that allow for clonal analysis, and the appearance of labelled cDCs and cDC clones in tissues will be followed over time in the steady-state or after induction of infection or inflammation. The dependence of cDC precursor activity on specific pathogen and damage sensing pathways will be assessed by loss-of-function experiments. The interactions of cDC precursors with their BM niche will be analysed in steady-state or inflammatory conditions by visualising the cells in situ. Finally, the consequences of demand-driven cDCpoiesis for immunity will be assessed. The results from this project will lead to a greater understanding of the influence of environmental signals on cDCpoiesis and may have applications in the design of better vaccines and immunotherapies.

Classical dendritic cells (cDCs) are leucocytes that play a key role in innate immunity as well as the initiation and regulation of T cell responses. cDCpoiesis starts with commitment of a bone marrow (BM) haematopoietic progenitor, known as the classical DC precursor (CDP), to the cDC lineage. CDPs then give rise to pre-cDCs that exit the BM via the blood and seed tissues to give rise to the two major types of fully-differentiated cDCs, the cDC1 and cDC2 subsets. The key parameters of cDCpoiesis are poorly understood. We propose to characterise the niche in which cDCs develop within the BM and to study how pre-cDCs seed tissues and establish local clones of differentiated cDC1 and cDC2. We further wish to ask how the activity of CDPs and pre-cDCs is altered following infection, inflammation or tissue damage. Finally, we want to know to what extent cDCpoiesis is affected by direct sensing of infection or cell damage by cDC precursors. All these objectives will be addressed in a mouse lineage tracing model in which cDC precursors are genetically labelled through the activity of a Cre recombinase driven by the Clec9a locus. These mice will be crossed to fluorescent protein reporter mice, including Confetti mice that allow for clonal analysis, and the appearance of labelled cDCs and cDC clones in tissues will be followed over time in the steady-state or after induction of infection or inflammation. The dependence of cDC precursor activity on specific pathogen and damage sensing pathways will be assessed by loss-of-function experiments. The interactions of cDC precursors with their BM niche will be analysed in steady-state or inflammatory conditions by visualising the cells in situ. Finally, the consequences of demand-driven cDCpoiesis for immunity will be assessed. The results from this project will lead to a greater understanding of the influence of environmental signals on cDCpoiesis and may have applications in the design of better vaccines and immunotherapies.

Max ERC Funding

2 500 000 €

Duration

Start date: 2018-09-01, End date: 2023-08-31

Project acronymDCSUBSET

ProjectCross-species characterisation of CD8alpha+ dendritic cells and their role in immune regulation

Researcher (PI)Caetano Maria Pacheco Pais Dos Reis E Sousa

Host Institution (HI)THE FRANCIS CRICK INSTITUTE LIMITED

Call DetailsAdvanced Grant (AdG), LS6, ERC-2010-AdG_20100317

SummaryDendritic cells (DC) are a heterogeneous family of leucocytes with important functions in immunity. Little is known about the role of distinct DC subtypes in vivo. In the mouse, a subset known as CD8alpha+ DC has been argued to represent a discrete DC lineage with specialised properties. These include a superior capacity for presenting exogenous antigens to CD8+ and CD4+ T cells, which makes CD8alpha+ DC an attractive target in vaccination and tolerisation. However, it is unclear whether CD8alpha+ DC fulfill unique and non-redundant roles in the immune system. In addition, the reported restriction of CD8alpha+ DC to thymus and secondary lymphoid organs is hard to reconcile with their documented capacity to act as presenting cells for antigens outside those organs. Finally, CD8alpha+ DC have not been identified in humans, greatly restricting their use in immunotherapy. In this proposal, we exploit the recent finding that DNGR-1 (CLEC9A) acts as a new and specific marker for the CD8alpha+ lineage to address these issues. We will generate DNGR-1-Cre mice as a universal tool to manipulate gene expression in the subset. We will use such mice to render CD8alpha+ DC sensitive to toxic proteins that permit constitutive or transient ablation of the subset for functional studies. DNGR-1-Cre mice will further be used to express fluorescent proteins in CD8alpha+ DC, allowing tracing of the lineage in vivo, both in lymphoid and non-lymphoid organs. Finally, we will use the DNGR-1 marker to identify and characterise putative CD8alpha+ DC equivalents in humans. The results from this proposal will illuminate the function of CD8alpha+ DC across species and open the door for using this intriguing DC subset in immunotherapy of cancer, infectious and autoimmune diseases.

Dendritic cells (DC) are a heterogeneous family of leucocytes with important functions in immunity. Little is known about the role of distinct DC subtypes in vivo. In the mouse, a subset known as CD8alpha+ DC has been argued to represent a discrete DC lineage with specialised properties. These include a superior capacity for presenting exogenous antigens to CD8+ and CD4+ T cells, which makes CD8alpha+ DC an attractive target in vaccination and tolerisation. However, it is unclear whether CD8alpha+ DC fulfill unique and non-redundant roles in the immune system. In addition, the reported restriction of CD8alpha+ DC to thymus and secondary lymphoid organs is hard to reconcile with their documented capacity to act as presenting cells for antigens outside those organs. Finally, CD8alpha+ DC have not been identified in humans, greatly restricting their use in immunotherapy. In this proposal, we exploit the recent finding that DNGR-1 (CLEC9A) acts as a new and specific marker for the CD8alpha+ lineage to address these issues. We will generate DNGR-1-Cre mice as a universal tool to manipulate gene expression in the subset. We will use such mice to render CD8alpha+ DC sensitive to toxic proteins that permit constitutive or transient ablation of the subset for functional studies. DNGR-1-Cre mice will further be used to express fluorescent proteins in CD8alpha+ DC, allowing tracing of the lineage in vivo, both in lymphoid and non-lymphoid organs. Finally, we will use the DNGR-1 marker to identify and characterise putative CD8alpha+ DC equivalents in humans. The results from this proposal will illuminate the function of CD8alpha+ DC across species and open the door for using this intriguing DC subset in immunotherapy of cancer, infectious and autoimmune diseases.

Max ERC Funding

2 499 998 €

Duration

Start date: 2011-09-01, End date: 2017-08-31

Project acronymDUT-signal

ProjectdUTPase Signalling: from Phage to Eukaryotes

Researcher (PI)Jose Rafael Penades Casanova

Host Institution (HI)UNIVERSITY OF GLASGOW

Call DetailsAdvanced Grant (AdG), LS6, ERC-2014-ADG

SummarydUTPases (DUTs) are enzymes that regulate cellular dUTP levels to prevent the misincorporation of uracil into DNA. Recently however, DUTs have been involved in the control of relevant cellular processes. How these regulatory functions are controlled remains unsolved. The recent elucidation of the mechanistic role of DUTs in the transfer of staphylococcal pathogenicity islands (SaPIs) by our group has revealed an entirely novel and surprising strategy involving DUTs in signalling. Namely, we have demonstrated that in addition to the 5 classical domains present in all the trimeric DUTs, staphylococcal phage-encoded DUT proteins possess an extra region (Motif VI) involved in SaPI de-repression by binding to the SaPI-encoded repressor (Stl). Although this domain is necessary, it does not suffice to induce the SaPI cycle. Unexpectedly, the strongly conserved DUT motif V is also inherently involved in mediating de-repression. Crystallographic and mutagenic analyses have demonstrated that binding to dUTP orders the C-terminal motif V of phage-encoded DUTs, potentially rendering these proteins in the conformation required for SaPI de-repression. In contrast, conversion into the apo state conformation by the hydrolysis of the bound dUTP disorders motif V and generates a protein that is unable to induce the SaPI cycle. Analogously, previous work demonstrated that the trimeric rat DUT interacts with the transcriptional factor PPARα, an interaction that depends on an “extra” N-terminal motif VI present in the DUT protein and requires the C-terminal domain contribution, strongly supporting in general the mechanism involving DUTs in signalling. In summary, our results suggest that DUTs define a widespread family of signalling molecules that acts analogously to eukaryotic G-proteins. This project stems from this ground-breaking result, and will investigate the biological role of DUTs as signalling molecules, opening up the possibility to establish dUTP as a new second messenger.

dUTPases (DUTs) are enzymes that regulate cellular dUTP levels to prevent the misincorporation of uracil into DNA. Recently however, DUTs have been involved in the control of relevant cellular processes. How these regulatory functions are controlled remains unsolved. The recent elucidation of the mechanistic role of DUTs in the transfer of staphylococcal pathogenicity islands (SaPIs) by our group has revealed an entirely novel and surprising strategy involving DUTs in signalling. Namely, we have demonstrated that in addition to the 5 classical domains present in all the trimeric DUTs, staphylococcal phage-encoded DUT proteins possess an extra region (Motif VI) involved in SaPI de-repression by binding to the SaPI-encoded repressor (Stl). Although this domain is necessary, it does not suffice to induce the SaPI cycle. Unexpectedly, the strongly conserved DUT motif V is also inherently involved in mediating de-repression. Crystallographic and mutagenic analyses have demonstrated that binding to dUTP orders the C-terminal motif V of phage-encoded DUTs, potentially rendering these proteins in the conformation required for SaPI de-repression. In contrast, conversion into the apo state conformation by the hydrolysis of the bound dUTP disorders motif V and generates a protein that is unable to induce the SaPI cycle. Analogously, previous work demonstrated that the trimeric rat DUT interacts with the transcriptional factor PPARα, an interaction that depends on an “extra” N-terminal motif VI present in the DUT protein and requires the C-terminal domain contribution, strongly supporting in general the mechanism involving DUTs in signalling. In summary, our results suggest that DUTs define a widespread family of signalling molecules that acts analogously to eukaryotic G-proteins. This project stems from this ground-breaking result, and will investigate the biological role of DUTs as signalling molecules, opening up the possibility to establish dUTP as a new second messenger.

Max ERC Funding

2 246 192 €

Duration

Start date: 2015-12-01, End date: 2020-11-30

Project acronymE-T1IFNs

ProjectElaboration of the type I interferonopathies

Researcher (PI)Yanick Joseph CROW

Host Institution (HI)THE UNIVERSITY OF EDINBURGH

Call DetailsAdvanced Grant (AdG), LS6, ERC-2017-ADG

SummaryType I interferons represent both key molecules in anti-viral defence and mediators of inflammatory disease, so that the induction, transmission and resolution of the interferon response are tightly regulated - balancing protection against infection versus the risk of immunopathology. Monogenic type I interferonopathies (T1IFNs), and related ‘complex’ phenotypes such as systemic lupus erythematosus and dermatomyositis, represent examples of a disturbance of the homeostatic control of this system, where a constitutive upregulation of type I interferon activity is considered directly relevant to pathology.
Set against the absence of a routine assay in clinical medicine for the detection of upregulated type I interferon, the current application addresses major questions in the developing T1IFN field. Analogous to other screening strategies (e.g. using mouse ENU mutagenesis or yeast gene deletion series), we have established a pipeline for the systematic identification of human mutant states predisposing to upregulated type I interferon signalling. Such an approach will allow for the comprehensive definition of important themes in interferon biology, informing our understanding of anti-viral signalling and self-non-self discrimination. Furthermore, these studies will have direct translational benefit - since the identification of a phenotype as a T1IFN implies the possibility of therapy to reduce type I interferon levels and / or block interferon signalling.

Type I interferons represent both key molecules in anti-viral defence and mediators of inflammatory disease, so that the induction, transmission and resolution of the interferon response are tightly regulated - balancing protection against infection versus the risk of immunopathology. Monogenic type I interferonopathies (T1IFNs), and related ‘complex’ phenotypes such as systemic lupus erythematosus and dermatomyositis, represent examples of a disturbance of the homeostatic control of this system, where a constitutive upregulation of type I interferon activity is considered directly relevant to pathology.
Set against the absence of a routine assay in clinical medicine for the detection of upregulated type I interferon, the current application addresses major questions in the developing T1IFN field. Analogous to other screening strategies (e.g. using mouse ENU mutagenesis or yeast gene deletion series), we have established a pipeline for the systematic identification of human mutant states predisposing to upregulated type I interferon signalling. Such an approach will allow for the comprehensive definition of important themes in interferon biology, informing our understanding of anti-viral signalling and self-non-self discrimination. Furthermore, these studies will have direct translational benefit - since the identification of a phenotype as a T1IFN implies the possibility of therapy to reduce type I interferon levels and / or block interferon signalling.

Max ERC Funding

2 418 800 €

Duration

Start date: 2018-11-01, End date: 2023-10-31

Project acronymELFBAD

ProjectL-form bacteria, biotechnology and disease

Researcher (PI)Jeffery Errington

Host Institution (HI)UNIVERSITY OF NEWCASTLE UPON TYNE

Call DetailsAdvanced Grant (AdG), LS6, ERC-2014-ADG

SummaryDespite the clear importance and multiple functions of the bacterial cell wall, many bacteria appear to be able to switch into a cell wall deficient or “L-form” state. L-forms are very heterogeneous in size and shape and generally require osmotic stabilisers, such as 0.5 M sucrose, for viability.
However, by lacking the requirement for a cell wall, L-forms are completely resistant to common cell wall antibiotics, such as β-lactams, and they are probably protected from some elements of innate immune recognition. L-forms are therefore of potential interest in relation to their possible involvement in human disease. They have often been reported in clinical specimens obtained from patients with recurrent or persistent infections or on long term prophylaxis with β-lactam antibiotics. Unfortunately, until recently, most of the work on L-forms had been done in the pre-molecular era, when it was difficult to characterise the L-forms and particularly to identify their origins and relationship with other resident pathogenic bacteria. Recently, several labs have revisited the L-form issue and started to apply modern molecular and cell biological methods.
The proposal is divided into three Themes:
• Improve our understanding of key features of the L-forms of our best characterised model system, B. subtilis, including both basic science and possible biotechnological applications.
• Extend our analysis of basic L-form biology into several diverse bacterial systems, of relevance to both biotechnology and infectious disease.
• Explore in detail the possible clinical relevance of L-forms, aiming to identify specific clinical situations in which they are relevant or, at least, to establish model systems in which the interactions between L-form and mammalian systems can be studied.

Despite the clear importance and multiple functions of the bacterial cell wall, many bacteria appear to be able to switch into a cell wall deficient or “L-form” state. L-forms are very heterogeneous in size and shape and generally require osmotic stabilisers, such as 0.5 M sucrose, for viability.
However, by lacking the requirement for a cell wall, L-forms are completely resistant to common cell wall antibiotics, such as β-lactams, and they are probably protected from some elements of innate immune recognition. L-forms are therefore of potential interest in relation to their possible involvement in human disease. They have often been reported in clinical specimens obtained from patients with recurrent or persistent infections or on long term prophylaxis with β-lactam antibiotics. Unfortunately, until recently, most of the work on L-forms had been done in the pre-molecular era, when it was difficult to characterise the L-forms and particularly to identify their origins and relationship with other resident pathogenic bacteria. Recently, several labs have revisited the L-form issue and started to apply modern molecular and cell biological methods.
The proposal is divided into three Themes:
• Improve our understanding of key features of the L-forms of our best characterised model system, B. subtilis, including both basic science and possible biotechnological applications.
• Extend our analysis of basic L-form biology into several diverse bacterial systems, of relevance to both biotechnology and infectious disease.
• Explore in detail the possible clinical relevance of L-forms, aiming to identify specific clinical situations in which they are relevant or, at least, to establish model systems in which the interactions between L-form and mammalian systems can be studied.

Max ERC Funding

2 428 621 €

Duration

Start date: 2015-10-01, End date: 2020-09-30

Project acronymGENINVADE

ProjectParasite population genomics and functional studies towards development of a blood stage malaria vaccine

Researcher (PI)David Joseph Conway

Host Institution (HI)LONDON SCHOOL OF HYGIENE AND TROPICAL MEDICINE

Call DetailsAdvanced Grant (AdG), LS6, ERC-2011-ADG_20110310

SummaryAn effective malaria vaccine is needed, particularly against P. falciparum as this species causes more human mortality than all other eukaryotic pathogens combined. An understanding of natural selection operating on parasites in local endemic populations can enable understanding of core molecular mechanisms of global relevance. The objectives are to
- Advance understanding of alternative pathways of erythrocyte invasion by malaria parasites
- Advance understanding of immune evasion by malaria parasites
- Identify optimal combinations of parasite proteins as malaria vaccine candidates
- Develop the interface between population genomic and functional studies of malaria parasites
The research programme will take an integrated approach to understanding pathogen adaptation, by designing experiments that are based on analysis at the molecular, functional, and population levels.
(i) Population genetic analyses of P. falciparum in sites of contrasting endemicity in West Africa, to finely determine signatures of selection with high-resolution throughout the genome, and help refine hypotheses on mechanisms used by merozoites to invade erythrocytes and evade acquired immune responses.
(ii) Experimental culture analysis of merozoite invasion into erythrocytes to identify the receptor-ligand interactions used by different parasite populations ex vivo. Novel receptor knockdown assays on cultured erythrocytes will be employed, and parasite adaptation experiments conducted to identify constraints on the use of alternative invasion pathways
(iii) Innovative approaches to select individual parasites and characterise cell tropism, transcript profiles, and genome sequences. This is aimed to validate population level findings and revolutionise approaches to genetics and phenotyping of parasites in the future. Candidate molecule discoveries will be taken forwards to receptor-ligand interaction assays, antibody inhibition and immuno-epidemiological studies.

An effective malaria vaccine is needed, particularly against P. falciparum as this species causes more human mortality than all other eukaryotic pathogens combined. An understanding of natural selection operating on parasites in local endemic populations can enable understanding of core molecular mechanisms of global relevance. The objectives are to
- Advance understanding of alternative pathways of erythrocyte invasion by malaria parasites
- Advance understanding of immune evasion by malaria parasites
- Identify optimal combinations of parasite proteins as malaria vaccine candidates
- Develop the interface between population genomic and functional studies of malaria parasites
The research programme will take an integrated approach to understanding pathogen adaptation, by designing experiments that are based on analysis at the molecular, functional, and population levels.
(i) Population genetic analyses of P. falciparum in sites of contrasting endemicity in West Africa, to finely determine signatures of selection with high-resolution throughout the genome, and help refine hypotheses on mechanisms used by merozoites to invade erythrocytes and evade acquired immune responses.
(ii) Experimental culture analysis of merozoite invasion into erythrocytes to identify the receptor-ligand interactions used by different parasite populations ex vivo. Novel receptor knockdown assays on cultured erythrocytes will be employed, and parasite adaptation experiments conducted to identify constraints on the use of alternative invasion pathways
(iii) Innovative approaches to select individual parasites and characterise cell tropism, transcript profiles, and genome sequences. This is aimed to validate population level findings and revolutionise approaches to genetics and phenotyping of parasites in the future. Candidate molecule discoveries will be taken forwards to receptor-ligand interaction assays, antibody inhibition and immuno-epidemiological studies.

Max ERC Funding

2 948 083 €

Duration

Start date: 2012-07-01, End date: 2017-06-30

Project acronymHIVINNATE

ProjectCharacterisation and Manipulation of Primate Lentiviral Interactions with Innate Immunity

Researcher (PI)Gregory John Towers

Host Institution (HI)UNIVERSITY COLLEGE LONDON

Call DetailsAdvanced Grant (AdG), LS6, ERC-2013-ADG

SummaryOur aim is to seek detailed molecular level understanding of the interactions between HIV-1 and innate immune sensors expressed in myeloid cells. We have demonstrated that HIV-1 replicates in primary human macrophages without triggering interferon production. However, by specific mutation of HIV-1 proteins or by manipulating interaction with host cofactors we can reveal the virus to innate immune receptors and activate an antiviral response leading to secretion of soluble type 1 interferon and cessation of replication. We propose to define the sensors and the details of the antiviral pathways that are activated in macrophages using proven RNA interference techniques reading out activation of innate immune responses by measurement of secreted interferon and induction of gene expression. We have also characterised small molecules that potently inhibit HIV-1 by revealing HIV-1 to innate immune sensors. In collaboration with crystallographers and medicinal chemists we aim to improve the potency and specificity of these drugs and to use them to study the anti-HIV-1 innate immune response. DC are sentinels of innate immunity and their infection induced maturation leads to interferon production and DC dependent T cell maturation that defines the nature and potency of the immune response. We will examine the effect of triggering innate responses in DC using HIV-1 mutants/drug treated wild type virus on allogeneic responses, by measurement of T cell proliferation and function and in an ex vivo CD8 T cell killing assays using peripheral blood CD8 cells from HIV‑1 infected patients. In this way we will uncover the molecular details of HIV-1’s interaction with innate immunity and discover how the virus replicates in primary immune cells without detection. This work will make a significant technical and intellectual contribution to an important emerging scientific field focusing on understanding and manipulating the complex relationship between HIV-1 and innate immunity.

Our aim is to seek detailed molecular level understanding of the interactions between HIV-1 and innate immune sensors expressed in myeloid cells. We have demonstrated that HIV-1 replicates in primary human macrophages without triggering interferon production. However, by specific mutation of HIV-1 proteins or by manipulating interaction with host cofactors we can reveal the virus to innate immune receptors and activate an antiviral response leading to secretion of soluble type 1 interferon and cessation of replication. We propose to define the sensors and the details of the antiviral pathways that are activated in macrophages using proven RNA interference techniques reading out activation of innate immune responses by measurement of secreted interferon and induction of gene expression. We have also characterised small molecules that potently inhibit HIV-1 by revealing HIV-1 to innate immune sensors. In collaboration with crystallographers and medicinal chemists we aim to improve the potency and specificity of these drugs and to use them to study the anti-HIV-1 innate immune response. DC are sentinels of innate immunity and their infection induced maturation leads to interferon production and DC dependent T cell maturation that defines the nature and potency of the immune response. We will examine the effect of triggering innate responses in DC using HIV-1 mutants/drug treated wild type virus on allogeneic responses, by measurement of T cell proliferation and function and in an ex vivo CD8 T cell killing assays using peripheral blood CD8 cells from HIV‑1 infected patients. In this way we will uncover the molecular details of HIV-1’s interaction with innate immunity and discover how the virus replicates in primary immune cells without detection. This work will make a significant technical and intellectual contribution to an important emerging scientific field focusing on understanding and manipulating the complex relationship between HIV-1 and innate immunity.

Host Institution (HI)THE CHANCELLOR, MASTERS AND SCHOLARS OF THE UNIVERSITY OF OXFORD

Call DetailsAdvanced Grant (AdG), LS6, ERC-2011-ADG_20110310

SummaryDefining the genetic basis of differential susceptibility to infectious diseases is of importance for understanding the evolution of human genetic diversity, for identifying critical molecular pathways in disease resistance, and for the design of novel intervention strategies such as more effective vaccines.
I propose to sequence the entire coding regions of all human genes in large numbers of cases of severe tuberculosis and fatal bacterial sepsis to identify variants that have a large impact on risk of developing severe tuberculosis or dying from sepsis. I shall then apply this exome sequencing approach to define the genetic basis of variable immune responsiveness in West Africans to hepatitis B vaccine, and to a new promising T cell-inducing vaccine, developed in my group, that targets the liver-stage of malaria.
This programme will benefit from unique collections of 10,000 disease cases and in-house expertise in vaccine design, bioinformatics and statistical genetics, and will take genetic investigation of common infectious disease to near the ultimate level of analysis by using large-scale next generation sequencing.

Defining the genetic basis of differential susceptibility to infectious diseases is of importance for understanding the evolution of human genetic diversity, for identifying critical molecular pathways in disease resistance, and for the design of novel intervention strategies such as more effective vaccines.
I propose to sequence the entire coding regions of all human genes in large numbers of cases of severe tuberculosis and fatal bacterial sepsis to identify variants that have a large impact on risk of developing severe tuberculosis or dying from sepsis. I shall then apply this exome sequencing approach to define the genetic basis of variable immune responsiveness in West Africans to hepatitis B vaccine, and to a new promising T cell-inducing vaccine, developed in my group, that targets the liver-stage of malaria.
This programme will benefit from unique collections of 10,000 disease cases and in-house expertise in vaccine design, bioinformatics and statistical genetics, and will take genetic investigation of common infectious disease to near the ultimate level of analysis by using large-scale next generation sequencing.